Cytomegalovirus (HCMV) is the most common human congenital infection worldwide and frequently causes multi-organ disease in immunocompromised hosts. In adults, the importance of HCMV-specific cell-mediated immune responses has been established for control of HCMV viremia and protection against disease development. However, the capability of young human infants to generate virus-specific cell-mediated immunity has not been well defined. We therefore propose to study the development of HCMV-specific:CD4+ and CD8+ T lymphocyte responses in infants experiencing primary infection. We hypothesize that virus-specific T lymphocyte responses will demonstrate qualitative and quantitative changes over time following primary infection. To address this hypothesis, we will characterize the magnitude and breadth of HCMV-specific CD8+ T cell responses in infants and children from acute through chronic infection. We will first use ELISPOT assays to identify the hierarchy of recognition of HCMV proteins. Epitopes within the most commonly recognized HCMV proteins and their HLA Class I-restricting elements will be defined. Tetramers representing commonly recognized epitopes will be made and used to characterize changes in HCMV epitope-specific CD8+ T cell frequencies over time. We will then characterize the cell surface phenotypes cellular turnover, and activation state of HCMV epitope-specific T cells using tetramer-staining, along with monoclonal antibodies to cell-surface antigens; functional properties (lytic function, chemokine/cytokine secretion) of HCMV epitope-specific CD8+ T cells will be evaluated. The frequencies and specificities of HCMV-specific CD4+ T cells will also be examined over time and correlated with HCMV-specific CD8+ T cell frequencies. Concurrent measurement of blood HCMV load will allow us to examine the relationship between HCMV-specific cell-mediated immune responses and blood HCMV load. Data from these studies will contribute to our understanding of age-related susceptibility to viral infections and will complement ongoing NIH-funded projects in our laboratory focused on examining the ability of young human infants to generate HIV-1 specific cell-mediated immune responses. The characterization of HCMV-specific cell-mediated immune responses and HCMV viral load over time following infection should help us to better understand virus-host interactions that contribute to the establishment of persistent viral infections and should aid in the development of improved prophylactic and therapeutic interventions for pediatric HCMV infection. [unreadable] [unreadable]